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Genome-Wide Association Study of Cleft Lip and Cleft Palate
Overview Cleft lip with or without cleft palate is a common birth defect in which tissues forming the lip do not join completely before birth. The lip forms between the fourth and seventh week of pregnancy, and during this time the tissues fail to bind causing an opening in the upper lip that is variable in size and severity. Later on in fetal development the roof of the mouth may also fail to join creating a similar absence of tissue in the palate. The condition, while occasionally occurring in sequence with other facial deformations, occurs as an isolated defect 70% of the time. The evidence for genetic linkage is strong, and studies have shown a familial recurrence rate 20-30 times greater than that of the general population. It had been previously determin ed that there is strong evidence in patients of European descent express an allele that disrupts the function of AP-2 binding site near the IRF6 gene. There are likely many more risk factors, and so a genome wide association study (GWAS) was performed to identify and confirm risk alleles. Oral Cleft GWAS Method: The formation of the International Cleft Consortium in 2007 allowed for the assembly of a 1591 case-parent trios from the USA, China, Philippines, Denmark, and Colombia among others. The cases of isolated, non-syndromic oral cleft we ascertained through treatment centers by members of the consortium. The cases were given physical examinations to identify any other deformities at play, and parents were interviewed about familial history of clefts, and other potentially contributing factors such as medical history, pregnancy history, and maternal exposure to potential risk factors. The DNA was collected from each member of the trio and sequenced, and the genetic variants were determined using SNP arrays. Results: In this GWAS it was found that multiple SNPs on chr. 8q24 and 4 SNPs in IRF6 showed genome wide significance. SNPs in two genes not previously associated with CL/P,ABCA4on chr. 1p22.1 andMAFBon chr. 20q12, were measured to have genome wide significance as well.MAFB,which is expressed in the mouth palatal shelf, experienced a rare missense mutation (MAFBH131Q) that was over-represented in the Filippino population, but was absent in other populations. ABCA4is a gene that encodes for transmembrane proteins, and mutations are noted in juvenile macular degeneration, however no previous evidence has indicated a role in clefting. Three more genes were identified to have some potential impact in clefting, but were not measured to be statistically significant to the whole genome. These genes includedPAX7 on chr. 1p36, VAX1 on 10q25.3 and NTN1 on 17p13. The SNP on chr. 8q24 sowing the strongest significance was rs987525. The odds ratio of rs987525 (A;G) was 2.57, while the odds ratio of rs987525 (A;A) was 6.05 suggesting that this SNP is a major genetic cause. It is also important to note that the significance of rs987525 was not comarable amongst all groups, and Europeans and South Americans showed a higher level of significance than Asians. There is a lower minor allele frequency among Asians, the cause of which was not determined. Replication in independent samples focused on 5 SNPs (rs987525 and 2 SNPs in MAFB and ABCA4). The attached table shows the relative significance of these genes between source populations. It is notable that families of European descent show greater significance for rs987525 on chr. 8q24 while families of Asian ancestry show a greater significance for MAFB and ABCA4. GWAS Significance This GWAS identified four SNPs near the genes MAFB and ABCA4 that were measured to be statistically significant in cause CL/P, and confirmed two previously identified regions (IRF6 and rs987525) to have genome-wide significance. Oral clefts are rarely life threatening and present minimal disease state beyond cosmetic deformity. However, the process of restoration includes sequential surgeries that carry significant dental and cosmetic complications. In underdeveloped regions of the world the prevalence of oral cleft is similar to that of developed nations, but the rate of surgical intervention is significantly decreased leading to increased risks of complications that come with oral deformities. Understanding the cause of genetic deformities is key to preventing them. At this point in time it seems that the more genetic studies done make the causes of these diseases increasingly convoluted, however consistent contributions to literature are key in helping us develop our understanding what causes deformities and how we could possibly inhibit these contributing factors. References: Beaty TH, Murray JC (2010): GWAS of cleft lip with and without cleft palate identifies risk varients near MAFB and ABCA4 Medline Plus; National Institutes of Health: Cleft Lip and Palate Rahimov F, Marazita ML (2008): Disruption of an AP-2alpha binding site in an IRF6 enhance is associated with cleft lip